Unlocking the PirAB Pandora's box: Genomic epidemiology and phage biocontrol of an emerging waterborne one health crisis.

Intensive aquaculture promotes the emergence of waterborne pathogens and the accumulation of antimicrobial resistance (AMR), challenging sustainable water management. Acute hepatopancreatic necrosis disease (AHPND), driven by PirAB-producing bacteria, severely threatens crustacean aquaculture. However, the broader environmental distribution of PirAB genes and feasible non-antibiotic control strategies remain insufficiently characterized. Here, we integrated large-scale comparative genomics with experimental phage validation to investigate the dissemination and biocontrol of PirAB-positive (PirAB+) bacteria. Screening 2.37 million bacterial genomes revealed that PirAB homologues span five phyla and 31 genera, extending beyond classical AHPND-associated Vibrio to aquatic, food-associated, and clinical microbiomes. PirAB+ Vibrio strains carried zoonotic virulence markers and elevated AMR burdens, highlighting their potential relevance to public health and water biosecurity. To support biocontrol, we constructed a genomic resource comprising 17,058 phage sequences associated with PirAB+ Vibrio, revealing extensive unexplored viral diversity, broad predicted host interactions, and diverse endolysins. We further isolated 11 phages from aquaculture waters using 55 Vibrio parahaemolyticus strains, selecting two lytic phages for cocktail formulation. In a Cherax quadricarinatus AHPND model, this cocktail achieved 89.3% survival, comparable to florfenicol, while promoting recovery of immune activity and hepatopancreatic tissue integrity. Overall, this study shows that PirAB genes are embedded within a broader network of virulence- and resistance-associated determinants, while providing a genome-guided phage framework for non-antibiotic AHPND control. These findings support phage-based strategies to reduce antibiotic dependence, mitigate AMR selection, and improve aquaculture water biosecurity.